Rationale: The tendency of tumor cells to gain resistance to multiple forms of therapy concurrently has long been a consistent barrier to effective treatment of disease. This effect significantly reduces the treatment options available to the clinician, and makes disease-free recovery difficult for the patient. The ability to discern particular mechanisms governing multi-modality resistance and exploit them, therefore, could result in more efficacious treatment strategies. We have previously shown that stress-induced transcription factor activation (e.g., AP-1) results in decreased response to therapeutic modalities. Furthermore, others have established a link between epigenetic regulation of gene expression (such as hypermethylation of gene promoters) and the malignant phenotype. Interestingly, there also appears to be a correlation between AP-1 and an epigenetic regulator of transcription, DNA methyltransferase. We attempted to delineate a mechanistic pathway involving stress-induced transcriptional events and methylation to identify potential molecular targets for reversing multi-modality resistance in tumor cells. Research Synopsis: Through collaboration with Dr. Douglas Spitz (previously of Washington University and recently of the University of Iowa), we obtained an oxidative stress-resistant fibroblast cell line with morphological and physiological characteristics of transformed cells. Among other treatments, these cells were significantly more resistant to hydrogen peroxide, cisplatin, etoposide, and heat-induced radiosensitization modalities. Exposure of the oxidative stress-resistant cells to the quintessential redox stressor, hydrogen peroxide, and chemotherapeutic agents concurrently with indomethacin resulted in a loss of the resistant phenotype. Interestingly, we also found that these stress-resistant cells also overexpressed AP-1 constitutively; indomethacin also inhibited the basal activity of this transcription factor (Cancer Res 61: 3486-3492). In pursuing a mechanism for this action, we found that additional cell lines made by Dr. Tom Curran (St. Jude Children's Hospital) to genetically overexpress the c-Fos constituent of AP-1 also exhibit resistance to several chemotherapeutic agents (e.g., cisplatin, etoposide) and show increased expression levels of the epigenetic regulator DNMT-1 (Kaushal et al, in preparation). Microarray analysis and subsequent Western blotting confirmed that Bag-1, an anti-apoptotic protein, was upregulated in c-Fos and DNMT-1 overexpressing cell lines, identifying the Bag family as a potential marker and prospective target for reversing the resistant phenotype. An ongoing collaboration with Drs. Andrew Feinberg and Bert Vogelstein (Johns Hopkins University), making use of microarray technology in DNMT-deficient cells, has yielded several prospective avenues of continuing this investigation (Gius et al, submitted).